Treatments for obesity remain limited and with low success. Current therapies do not account for known differences in individual obesity susceptibility, which might be key for developing successful treatments. Susceptibility and severity to obesity are linked to individual differences in spontaneous physical activity (SPA). Our long-term goal is to provide the biological basis for development of personalized obesity therapies. To do this, we will use a novel model of differential susceptibility to obesity, the hig activity (HA) and low activity (LA) rats. HA rats are obesity resistant and higher signaling by orexin peptides than LA rats. The orexins are key modulators of SPA and energy balance. Our overall goal is to define the mechanisms underlying orexin involvement in to SPA and their relevance to obesity susceptibility in the HA/LA rat model. A long-standing hypothesis about the orexins is their functional specialization, which proposes that orexin neurons in the lateral hypothalamus (LH) mediate reward, while neurons in the perifornical area (PeF) and dorsomedial hypothalamus (DMH) mediate arousal. Aim 1 will determine which orexin neuron subpopulations (LH or PeF/DMH) are more relevant for the HA/LA phenotype. We will knock- down and over-express orexins in these areas to test their necessity or sufficiency for SPA in. Next, we will focus on orexin signaling through rostral LH (rLH). This pathway mediates increases in SPA and may be key for expression of the HA phenotype. Aim 2 will determine the relevance of rLH orexin signaling to obesity susceptibility in HA/LA rats. We will determine how obesity affects orexin rLH signaling in HA/LA rats and if blocking orexin responses in rLH increases obesity susceptibility in HA rats. Next, we will study interactions between rLH and nucleus accumbens shell (NAcSh). NAcSh interacts with orexin neurons to affect feeding and SPA. Inhibition of NAcSh GABAergic efferents increases SPA caused by orexin injection in rLH in LA, but not in HA rats. Aim 3 will study NAcSh and rLH interactions in the HA/LA phenotype. We will determine if there is a combined effect of rLH and NAcSh orexin signaling in HA rats, whether feeding responses after manipulation of NAcSh are different between HA/LA rats and define the neuroanatomical connections between NAcSh and rLH orexin-responsive neurons. These studies will fill the gap in knowledge of orexin neural circuitry in mediating phenotypic differences between HA and LA rats, which will improve our understanding of brain mechanisms underlying individual obesity susceptibility and enhance therapeutic approaches to obesity.